Sinh tng hp interferon v interleukin bng cng ngh vi sinh

GVHD: T.S TRN THANH THU SVTH: PHM TH THU NGN CHUYN CNG NGH VI SINH

L do chn ti
Interleukin v interferon l hai loi thuc s dng trong liu php sinh hc iu tr ung th. Nh vo cng ngh vi sinh ti t hp to ra nhiu IFN v IL gi thnh thp p ng nhu s dng ca con ngi m ra mt tng lai ngy cng tt p cho loi ngi trong vic bo v v nng cao sc kho cng ng

Ni dung
I. Khi qut v interferon III. Khi qut v interleukin 1.khi nim IFN 1.Khi nim IL 2. lch s pht hin IFN 2.Interleukin-2
3. phn loi IFN 4. tnh cht sinh hc IFN 5. C ch hot ng IFN 6. cc yu t kich thich sinh IFN 7. sn xut IFN 2.1 lch s nghin cu 2.2 cu to IL-2 2.3 Hot tinh sinh hc IL-2 2.4 C ch hot ng IL-2 2.5 Sn xut IL-2

II. ng dng interferon

1. ng dng iu tr cho con ngi 2. ng dng trong th y

IV. ng dng interleukin V. Ti liu tham kho

I. Khi qut v interferon

1.Khi nim IFN l protein hoc glycoprotein c tc dng ngn cn s xm nhim ca virus hoc km hm pht trin khi u v c vai tr quan trng trong h thng min dch th nht.

2. lch s pht hin

IFN c pht hin u tin vo nm 1957, do Alick Isaacs ( Anh) v Jean Lindenman (thuy s ) khi th nghim nui cy cc t bo phi g

Hin nay, ngi ta bit n 22 gen nhiu loi t bo khc nhau ca c th c kh nng sn xut IFN nh i thc bo,t bo lympho,t bo NK..

3.PH N LOI

Typ 1 INF
Typ 2

IFN-

IFN- IFN-

3.IFN Typ1
L nhng cht trung gian p ng sm min dch sm bm sinh chng li cc nhim trng virut. Gm 2 nhm IFN- v IFN- + IFN- c khong 20 polypeptit lin quan n cu trc. Mi polypeptit c m ha bi cc gen ring r. Ngun t bo chnh sn xut IFN- l bch cu n nhn. IFN- l glycoprotein gm 2 chui ging nhau vi khi lng phn t 21-24 kDa c m ha bi cc gen khc nhau

IFN-

IFN-

 Chc
+ c ch s sao chp ca virut + c ch s sinh tng hp protein + lm tng kh nng tan ca cc t bo git t nhin + iu chnh s biu hin ca phn t MHC1 v c ch hon ton biu hin ca MHC2

nng sinh hc IFN typ1

C ch hot ng IFN typ I

3.2 Interferon typ II

l IFN- hay cn gi l IFN min dch v chng ch yu do t bo T hot ho to thnh nn thc cht cng l mt lymphokin

IFN-

 Chc nng sinh hc IFN typ II

+ l cht sinh min dch + kch thch hot ng mnh ca cc i thc bo + tng s hin din ca phn t MHC1 + kch thch s hot ha tiu hy cc t bo git t nhin t hn IFN typ I + tc dng trc tip ln t bo lympho T v B thc y nhanh qu trnh bit ha cc t bo CD4 to thnh TH1

C ch hot ng IFN-

4. Cc tnh cht sinh hc ca IFN

Rt nhy cm vi vi cc enzym phn gii protein Bn vng vi Ph axit, bn vng vi nhit 56oC vn gi nguyn hot tnh. c tnh khng nguyn yu C hot tnh khng nguyn yu C tnh c hiu loi C tc dng c ch khng c hiu vi cc loi virut v nhiu loi t bo ung th Gia IFN v cm ng nguyn sinh ra n khng c tnh c hiu cp

5. C CH HOT NG IFN
Khi mt t bo m b nhim bi virus, n gii phng IFN. IFN s khuch tn vo cc t bo xung quanh. Khi lin kt vi th th trn b mt ca cc t bo ln cn, IFN bt u sn xut mt loi protein c th ngn chn s tng hp cc protein virus. iu ny ngn cn s ly lan ca virus trong c th.

chng ta u bit mt t bo c th bnh thng. T botnbokhisa b mt nhin,tn g u phn t interferon cc phng Tip btvirus vi rngth vkt nhp vosn xut vmng ttinsp cnhnTuy n khibi K n, interferon s lin bch cu l tuyn t bo, pht ny ca kch thch cht. T Hy theo, c vn c da xm victh th trn th u khngv t th. ch virus iu D cc cac n s ra th bo bn c s xy geneth xm loi virut nguy himbo, tms s1 k h trong tuyn phng th khc nu nhng nhp vo trong t thy sao c tng cc bo protein sn sinh t s ngn chn nomt cch chnh xc v sta t bo u ng rac khi khngra protein khng thnhng n tV vy l chng bngvirus. squ cng. Mt khng virus cu c mnh virus ny. cu chp mnh kh cch cc qu trnh c mt dng tinmy trao i cht interferon. ta.Vng, cng ri t bo xm nhp cc t bo y v sn sinh ra ca th chng v sau tuyn phng th th hai s nhy vo tng tr xm nhp vo c t bo nh vic b nhanh v n gin nht to ra interferon cng nh s dng chng trong c th. trnh v hm nay, chng ti strnhv mt phn ca tuyn phng thhot hai - cn c bit n vi khc. Cng trong qu ni ny, gen interferon c kch th trong ht nhn ca t bo tn gi interferon - v tho lun s sn sinh ra n. ch. Click xem bn dch Phim c ch hot ng IFN

6.Cc yu t thng xuyn kch thich sinh IFN

Vi khun, virut,protein
Ni c t vi khun,di nguyn ..

7. sn xut interferon
Cng ngh sn xut interferon trong t bo vi khun E.coli gm:

To gen m ho IFN
Thu nhn cc loi ARN thng tin m ho cc loi IFN tng ng V d: thu nhn gen m ho IFN- + cn tch chit ARN thng tin m ho IFN- t t bo bch cu ngi

To vect ti t hp

+ Bng k thut phn t, gn gen m ho IFN ngi vo vect biu hin c ci bin t thc khun th phage + To cc vect phage ti t hp mang gen m ho IFN ngi

to dng vect

To dng t bo E.coli ti t hp mang gen m ho IFN

Bin np phage ti t hp vo t bo E.coli, to dng E.coli ti t hp. kim tra kh nng biu hin gen th bin np

ln men thu sinh khi v thu nhn IFN

Thc hin k thut ln men trong cc thit b ln men cc iu kin mi trng thch hp thu sinh khi t bo, tch chit IFN

Tinh sch v thu nhn IFN thnh phn

Thu nhn v tinh sch IFN Kim tra hot tnh v thm ph gia (NaCl, KH2PO4,dextran) c th gi hot tnh v bo qun lu di. th nghim lm sng kim tra tnh an ton ca sn phm Tinh ch, tit trng, nghin cu iu kin v ng gi

II. ng dng
1.ng dng iu tr ca Interferon i vi con ngi
IFN- v IFN- c s dng trong iu tr nhiu bnh do vi rt gy nn. Hin nay IFN- c s dng hiu qu trong iu tr vim gan C cp v mn; vim gan B mn; HIV. IFN- c s dng trong iu tr a hng cu IFN - l v kh hu hiu trong iu tr cm A/H5N1

IFN c s dng thnh cng iu tr cm (t 1972), chng virus Herpes trong ghp thn (1979), ung th xng, ung th v, u ty v u lympho bo (nm 1979 M vi t l cha khi t 40 -70%). Nm 1989 cng M, thuc c dng cho bnh nhn ung th bng quang, ung th v v u hc t. IFN c s dng hiu qu trong iu tr vim gan siu vi B-C mn tnh v c tc dng loi tr virus gy nhim, chuyn i huyt thanh, ci thin men gan v ngn nga ung th tin trin

Mt s sn phn ca IFN

2. ng dng trong th y
2.1 Interferon c s dng nh l t dc trong vaccine
I VI GIA SC I VI GIA CM

Interferon alpha v beta c s dng kt hp vi vaccine phng bnh l mm long mng, hi chng PRRS.. cho hiu qu cao.

Interferon gamma khi c dng chung vi khng nguyn c tc dng tng cng p ng khng th th cp duy tr nng cao trong mt thi gian di. Hn na, vic kt hp khng nguyn vi Interferon gamma lm gim liu s dng vaccine

2.2 Interferon dng chn on bnh

IFN - dng chun on: bnh lao b , Johns disease, Brucellosis (bnh do vi khun gy ra) Ngoi ra cn dng chn on bnh lao, bnh phong hi ngi. IFN- dng chn on bnh IBR

2.3 Interferon dng trong phng, tr bnh cho gia sc, gia cm S dng IFN- trong vic phng, tr bnh cho gia sc, gia cm l mt gii php thay th khng sinh an ton- hiu qu cao. IFN- c dng iu tr cc bnh do virus gy ra trn gia sc, gia cm nh: bnh vim ng h hp, bnh vim v, PRRS, TGEV, cc bnh cm, Marek, Gumboro, vim gan B; Ngoi ra cn h tr iu tr vi khng sinh mang li hiu qu cao, hn ch hin tng nhn thuc ca cc vi khun gy bnh. => Tuy nhin, vic ng dng sn phm ny sang lnh vc th y b hn ch v vn gi thnh sn phm

III.KHI QUT V INTERLEUKIN

1.Khi nim:
Interleukin l mt h gm nhiu protein, glycoprotein xytokin c vai tr quan trng trong h thng min dch ca t bo v c th.

INTERLEUKIN-21

 Ngy nay, cc nh khoa hc pht hin hng chc loi interleukin khc nhau nh IL-1,IL-2 Chc nng ca nhiu loi interleukin trong h thng min dch c nghin cu tng i k nh: IL-1 c vai tr kch thch s tng sinh ca cc t bo min dch T v B, IL-4 c vai tr kch thch cc t bo B v hot ho s to thnh tng bo,gy hin tng tit khng th IgE. IL-2 l loi c tm ra u tin v hin c nhiu ng dng trng iu tri bnh

2 Interleukin-2
2.1 lch s pht hin
c

pht hin ln u tin nm 1975

Nhn Interleukin 2 cu trc tinh th

2.2 cu to IL-2
L mt glycoprotein gm 153 axit amin, khi lng 15kDa,gm bn chui peptit xon kiu Gen m ho IL-2 nm trn nhim sc th s 10,gen m ho IL-2 c tch dng nm 1983

2.3 hot tnh sinh hc

2.4 c ch hot ng IL-2

t bo lympho T hot tnh Kch hot t bo lympho T
t bo l cht th ng t bo lympho T tng sinh

t bo phn nhnh

Kch hot cc t bo vo qu trnh cht

2.5 sn xut IL-2 bng chng vi sinh vt ti t hp

1
To on gen m ho

Tch chit ARN t cc t bo l lch ca ngi bng

cc k thut sinh hc phn t Nhn on cADN m ho IL-2 t ARN thng tin vi cc cp mi c hiu thu nhn c on cADN m ho IL-2

To vect ti t hp

s dng k thut phn t ni on cADN m ho IL2 vo vect tch dng. C th s dng vect pCR 2.1TA, to vect ti t hp pCR 2.1-IL-2

bin np vect ti t hp vo t bo E.coli DH5

s dng k thut sc nhit hoc in xung to dng vi khun E.coli DH5 mang gen m ho IL-2

Hon thin gen m ho IL-2

Gii trnh t kim tra on gen m ho IL-2

Thit k mi gy t bin mt im glycosyl ho v nhn gen vi thit b PCR to c gen m ho IL-2 ca ngi mt im

glycosyl ho

To dng t bo vi khun E.coli BL 21 c kh nng biu hin gen IL-2

Bng k thut phn t thng dng,chuyn on IL-2 t vect pCR 2.1-TA sang vect biu hin pET32a (+) nh s l vi cp emzym gii hn Not I v Nco I. Sau ,bin np vect biu hin pET32a (+) mang gen IL-2 vo t bo vi khun E.coli BL21

thc hin qu trnh ln men v thu nhn IL-2

Cc chng vi khun E.coli BL21 mang gen m ho IL-2 c ln men trong thit b ln men mi trng LBA c b sung( nng cui 100mg/l) 370oC sau 3 gi cm ng bng IPTG. Thu protein tan trong dch ni bng phng php li tm, thu nhn protein khng tan bng phng php ph t bo

Tch v tinh sch IL-2

Thu nhn IL-2 t kt qu ln men Tch v tinh sch IL-2 bng sc k i lc Xc nh tnh an ton ca IL-2,th nghim lm sng, nghin cu cch bo qun v ng gi. => 1/2011 IL-2 c sn xut thnh cng vit nam

1/2011 IL-2 c sn xut thnh cng vit nam

S
t.

n xut IL-2 trong iu kin GMP ti Cng ty Vc-xin v Sinh phm s 1, B Y

IV. ng dng interleukin

IL-2 c s dng trong iu tr nhiu loi ung th nh: ung th bn quan, ung th thn.. Hin nay, c quan qun l dc phm v thc phm m (US FDA) cho php s dng IL-2 trong iu tr ung th hc t (melanoma) v ung th c tnh.cc th nghim sn xut v s dng IL-2 trong iu tr nhiu loi bnh ung th khc nhau ang c thc hin nhiu nc

ng dng ca IL trong chn on:

nh hng chn on trong cc bnh nhim D bo thi ghp tng nh hng chn on trong cc bnh t min.

Mt s thnh tu mi IL
Mi y cc chuyn gia thuc H Thomas Jefferson va pht hin ra interleukin-7 c kh nng nh bt mt s dng HIV ra khi ni n np. Hin nay IL-8 c dng iu tr bnh vy nn

Kt lun
IFN v IL l mt protein c bit c kh nng chng virut,chng t bo ung th v tng cng h min dch. IFN v IL c vi tr quan trng trong h thng min dch ca con ngi. Ngy nay, bng k thut di truyn c th sn xut IFN v IL vi s lng ln nn h gi thnh

Ti liu tham kho

1.C s cng ngh vi sinh vt v ng dng. NXB GD VIT NAM 2. http://vi.wikipedia.org/wiki/Interferon
3. http://elearning.lrc-hueuni.edu.vn/coe/mod/resource/view.php?id=1490 4.http://www.vast.ac.vn/index.php?option=com_content&view=article&id=1 053&catid=37%3Atin-khcn-trong-nc-&Itemid=34&lang=vi 5.http://thuviensinhhoc.com/index.php?option=com_content&view=section& id=14&Itemid=419